Methods and systems for measuring ascorbic acid

Abstract

Disclosed are methods and systems using liquid chromatography/tandem mass spectrometry (LC-MS/MS and 2D-LC-MS/MS) for the analysis of ascorbic acid in biological samples.

What is claimed is: 
     
         1 . A method for determining the presence or amount of ascorbic acid in a sample by tandem mass spectrometry, comprising:
 (a) generating a fragment ion from the ascorbic acid of with a mass to charge ratio (m/z) of about 115 by in-source fragmentation;   (b) generating one or more product ions of the ascorbic acid fragment ion by tandem mass spectrometry;   (c) detecting the presence or amount of one or more of the ascorbic acid fragment ion generated in step (a) or the one or more product ions of step (b) or both; and   (d) relating the detected ions to the presence or amount of the ascorbic acid in the sample.   
     
     
         2 . The method of  claim 1 , wherein the sample is subjected to a purification step prior to the initial fragmentation step (a). 
     
     
         3 . The method of  claim 2 , wherein the purification step comprises chromatography and/or precipitation of proteins. 
     
     
         4 . The method of  claim 3 , wherein the chromatography comprises high performance liquid chromatography (HPLC). 
     
     
         5 . The method of  claim 3 , wherein the chromatography comprises analytical liquid chromatography. 
     
     
         6 . The method of  claim 4 , wherein the sample is heated at the MS/MS interface. 
     
     
         7 . The method of  claim 1 , further comprising at least one of liquid-liquid extraction of the sample or dilution of the sample prior to mass spectrometry. 
     
     
         8 . The method of  claim 1 , wherein the product ions comprise ions having a mass/charge ratio (m/z) of about 87 and 59. 
     
     
         9 . The method of  claim 1 , wherein the fragment ion comprises 3,4-dihyroxyfuran-2(5H)-one. 
     
     
         10 . The method of  claim 1 , wherein the product ions comprise at least one of (Z)-prop-1-ene-1,2,3-triol diradical or (Z)-ethene-1,2-diol radical. 
     
     
         11 . The method of  claim 1 , further comprising detection of ascorbic acid over a range of from about 0.05 mg/dL to about 5 mg/dL. 
     
     
         12 . The method of  claim 1 , further comprising providing a biological sample believed to contain ascorbic acid. 
     
     
         13 . The method of  claim 1 , wherein the tandem MS/MS spectrometer is operated in an atmospheric pressure chemical ionization (APCI) mode. 
     
     
         14 . A system for determining the presence or amount of ascorbic acid in a test sample, the system comprising:
 a station for providing a test sample suspected of containing ascorbic acid;   a station for in-source fragmentation of the ascorbic acid to generate an ascorbic acid fragment ion; and   a station for mass spectrometry to generate product ions from the ascorbic acid fragment ion; and   a station to analyze the mass spectrum to determine the presence or amount of ascorbic acid in the test sample.   
     
     
         15 . The system of  claim 14 , further comprising a station for partially purifying the ascorbic acid from other components in the sample. 
     
     
         16 . The system of  claim 14 , further comprising a station for chromatographically separating the ascorbic acid from other components in the sample. 
     
     
         17 . The system of  claim 14 , wherein at least one of the stations is controlled by a computer.